1. Field of the Invention
The invention relates to a system for collecting a biological sample, a method for collection of a biological sample, a kit, and a solid matrix for depositing a biological sample.
2. Description of Related Art
The analysis of biological material has long been of paramount importance in the diagnosis and treatment of disease, in food and environmental analysis and in forensic investigations, in particular using histological and pathological techniques. Recent technological advances have broadened the scope of such investigations by facilitating analysis of nucleic acids and proteins, which has opened up a large number of further possibilities. Gene activity can, for example, be determined directly by analysis of RNA, in particular the messenger RNA (mRNA) in cells. Quantitative analysis of transcription templates (mRNA templates) in cells by means of modern molecular biological methods, such as real time reverse transcriptase polymerase chain reaction (real time RT-PCR) or gene expression chip analysis enables, for example the identification of incorrectly expressed genes, so that, for example, metabolic disorders, infections or the presence of cancer can be detected. The analysis of DNA from cells by molecular biological methods, such as PCR (polymerase chain reaction), RFLP (restriction fragment length polymorphism), ALFP (amplified fragment length polymorphism) or sequencing permits, for example, the detection of genetic defects or the determination of the HLA (human leukocyte antigen) type as well as other genetic markers. The analysis of genomic DNA and RNA is also used as direct evidence for infectious agents, such as viruses, bacteria, and the like.
As the benefits of being able to analyse certain components of biological samples, for example nucleic acids or proteins, have become known, and as the analyses themselves have become more accurate and more accessible, such analyses have become important and frequently used tools available not only to the medical and veterinary professions, but also in a wide range of other areas, such as in the analysis of forensic materials, pharmaceutical products and intermediates, foods and environmental materials. In many of these areas it is important to maintain the integrity of the molecular structure of a sample.
Current procedures for the collection, handling and/or transport of biological samples such as blood, saliva or cells employ solid matrices such as cellulose- or cotton-based papers or swabs which differ significantly in their constituents and size.
For buccal or vaginal smears, swabs with a head made of cellulose, cotton or polymer fibres are frequently used. Swabs with heads made from cotton or synthetic fibre and swabs with ejectable paper heads, for example, are available from various commercial suppliers. After collection of the biological sample with such swabs, the swab is usually dried, stored and transported in a dried form, or stored and transported in a medium containing nutrients, antibiotics or other preservatives.
In order to recover the biological sample from the swab, it is generally necessary to wash the sample away from the swab support material. The washing away of the sample, which is often done by contacting the swab with a lysis reagent, is generally inefficient, incomplete and cumbersome. To maximise the amount of sample recovered, the support material of the swab head needs first to be separated from its shaft. Depending on the type of swab, this is accomplished by hand, by cutting with scissors, with a scalpel or a razor blade, or by bending or ejection. Separation processes involving cutting require the cutting device to be thoroughly cleaned and sterilised or disposed of after each use to avoid cross-contamination. Cross-contamination is a problem for all analytical methods, but particularly when working with samples destined for analysis of the nucleic acids, especially when employing an amplification process such as the polymerase chain reaction (PCR). Cross-contamination can, in the worst case, lead to distortion or even total falsification of the analysis results. The cutting process also represents a significant risk of injury to the operator, which can result in incorporation of foreign, potentially infectious material.
Ejectable swabs, where the head of the swab is separated from the shaft by bending or pressing the swab shaft, represent an expensive alternative.
Both types of swab have in common that the separated swab head carrying the biological material necessarily remains in the sample tube after separation from the swab shaft. The separated swab head then absorbs the transportation and/or storage solution, decreasing the accessible volume in the tube and hindering an efficient recovery of the sample with a minimal amount of liquid. Since often only very small amounts of sample are concerned, often, for example, on a nanogram scale, it is desirable to avoid as far as possible any loss or over-dilution of the sample.
Other biological samples such as blood are generally collected and stored using papers or cards, which can be treated or untreated. Treated papers are usually treated with several different agents which inactivate pathogens and prevent microbial growth and DNA degradation. In order to separate target biomolecules it is necessary to punch out smaller pieces of paper or card from the paper carrying the dried sample material. This process requires the cleaning and sterilisation of the punching device after each punching action in order to avoid cross-contamination. Disposable punching equipment is commercially available, but is cumbersome and expensive. In addition, the small weight of the paper punch together with static electricity and normal air movement make it difficult to handle the punch and to transfer it to the bottom of the sample preparation vessel.
Furthermore, for optimal yield of the target biomolecule it is necessary to cut the support material into small pieces before introducing it into the process to recover the biological sample. The cutting process is cumbersome and difficult to automate and represents another potential source of cross-contamination and risk of injury of the operator. Not cutting the support material results in reduced yields of recovered biological sample and compromised sensitivity in the downstream application, such as analysis of the sample.
After transfer of the solid matrix into the recovery reagent, which is usually a lysis reagent, the target biomolecules are usually eluted from the solid support. The elution of target biomolecules from the solid matrix is inefficient, cumbersome and tends to suffer from retention of a proportion of the biomolecules on the solid matrix. Elution can be carried out chemically, physically or enzymatically, or by a combination thereof. For further isolation of the target biomolecule it is necessary to separate the solid matrix from the liquid containing the biomolecule. This is done by removal of the liquid by pipetting, or by removal of the solid matrix. Both processes are inefficient and can lead to loss of sample material unless large quantities of solvent are used. They are also difficult to automate, because the solid matrix can interfere with a liquid handling system. It is also difficult to design an automated system that is able to remove blood spots or swabs reliably from a vessel. Another problem is the amount of liquid that remains trapped in the solid matrix. Since this liquid comprises the target biomolecules, its removal with the solid matrix causes decreased yield and sensitivity.
A number of different solid matrices in the form of a blood card or swabs, for example swabs with heads made from cellulose, cotton, Dacron® or other polymeric fibres is known and commercially available. There is also a choice of several recovery and purification methods. There is, however, a lack of a standard procedure and system for collection, storage, stabilisation and purification of biological samples. This makes it necessary to carry out a lot of optimisation work in order to optimise the yield and performance of a given combination of support material and sample preparation method.
WO 01/60517 describes a method for taking a blood sample using a receptacle for receiving a sample, preferably blood, containing a solution for stabilising nucleic acids and a solid phase capable of binding nucleic acids. The sample, once transferred to the vessel, subsequently needs to be removed from the solid phase by repeated washing. Furthermore, the collection of the sample requires the use of a cannula to transfer the liquid sample from its source into the receptacle, which has been placed under a low pressure. This prevents the use of the receptacle for small amounts of sample, for example blood from a finger prick or a heel prick from an infant, and for samples which cannot easily be transferred via cannula, or only with some discomfort for the sample donor, such as saliva, urine or cerebrospinal fluid. The receptacle is also not suitable for solid or semi-solid samples since a solid or semi-solid sample does not mix sufficiently with the stabilising solution in the presence of the solid phase.
EP 819 696 A2 describes a method of isolating nucleic acid from a biological sample and proposes the use of a nucleic acid-binding solid phase, preferably silica, which is mixed with the sample together with a chaotropic substance. The method requires a large number of washing, drying and elution steps in order to first isolate the solid phase with the thereto-bound nucleic acid from the solution and then isolate the nucleic acids from the solid phase. Furthermore, too large an amount of silica can be saturating, such that beyond a given amount of silica no further nucleic acid is obtained from the sample.
WO 02/072870 A2 describes a method and a device for storing genetic material. The head portion of the device is composed of a solid matrix for adsorbing genetic material and a preserving mechanism to protect the genetic material from degradation. The device can, however, only be used for liquid samples. Furthermore, the solid matrix must be cut into smaller portions in order to carry out analysis of the genetic material, resulting in loss of material and risk for the operator.
US 2006/0099567 and US 2005/0276728 propose a storage device for biological materials comprising dissolvable or dissociable matrix material which coats a sample well in a sample plate. The matrix material coating together with the sample is dried and subsequently rehydrated for sample recovery. The proposed system is particularly intended for use in high throughput systems. The biological sample is, however, first collected using conventional techniques and as such the proposed system does not address the above-mentioned problems associated with collection of the sample and recovery of the sample from the collecting device. In addition, these documents are silent about the benefits of providing a sample system with means for dissolving the matrix material in the presence of a chaotropic salt.